Heparin Interaction with the Primed Polymorphonuclear Leukocyte CD11b Induces Apoptosis and Prevents Cell Activation

Heparin is known to have anti-inflammatory effects, yet the mechanisms are not completely understood. In this study, we tested the hypothesis that heparin has a direct effect on activated polymorphonuclear leukocytes (PMNLs), changing their activation state, and can explain its anti-inflammatory effect. To test our hypothesis, we designed both in vitro and ex vivo studies to elucidate the mechanism by which heparin modulates PMNL functions and therefore the inflammatory response. We specifically tested the hypothesis that priming of PMNLs renders them more susceptible to heparin. Amplified levels of CD11b and increased rate of superoxide release manifested PMNL priming. Increase in cell priming resulted in a dose-dependent increase in heparin binding to PMNLs followed by augmented apoptosis. Blocking antibodies to CD11b inhibited heparin binding and abolished the apoptotic response. Moreover, heparin caused a significant dose-dependent decrease in the rate of superoxide release from PMNLs, which was blunted by blocking antibodies to CD11b. Altogether, this study shows that the interaction of heparin with the PMNL CD11b results in cell apoptosis and explains heparin’s anti-inflammatory effects

Primed polymorphonuclear leukocytes from hemodialysis patients enhance monocyte transendothelial migration

Increased counts and priming of peripheral polymorphonuclear leukocytes (PMNLs) are associated with future or ongoing atherosclerosis; however, the role of PMNLs in enhancing monocyte transendothelial migration is still unclear. Our aims were to examine endothelial and monocyte activation, transmigration, and posttransmigration activation induced ex vivo by in vivo primed PMNLs and the effect of antioxidants on the activation. A unique ex vivo coculture system of three cell types was developed in this study, enabling interactions among the following: primary human umbilical vein endothelial cells (HUVECs), monocytes (THP-1 cell line), and in vivo primed PMNLs from hemodialysis (HD) patients and healthy control (HC) subjects. The interactions among these cells were examined, and an intervention with superoxide dismutase and catalase was performed. Preexposed HUVECs to HD/HC PMNLs showed a significant monocyte transmigration yield, 120–170% above HCs. Monocyte exposure to HD PMNLs induced pre- and posttransmigration activation. When the three cell types were cocultivated at the same time, monocyte chemoattractant protein-1 protein levels released from HUVECs, and activation markers on HUVECs [CD54 and chemokine (C-X3-C motif) ligand 1] and monocytes [chemokine (C-X3-C) receptor 1 and chemokine (C-C motif) receptor 2] were increased. Monocyte transmigration yield decreased to 70% (compared with HC subjects) due to adherence and accumulation of monocytes to HUVECs. When superoxide dismutase and catalase were used, reduced HUVEC and monocyte activation markers brought the transmigration yields to control levels and abolished accumulation of monocytes, emphasizing the role of superoxide in this process. We conclude that peripheral primed PMNLs play a pivotal role in enhancing monocyte transendotelial migration, the hallmark of the atherosclerotic process. Primed PMNLs can be used as a mediator and a biomarker of atherosclerosis even before plaque formation.NEW &amp; NOTEWORTHY Primed polymorphonuclear leukocytes are key mediators in monocyte transendothelial migration, a new understanding of the initiation of endothelial dysfunction and monocyte activation, transmigration, and accumulation in the subendothelial layer.</jats:p

Heparin Interaction with the Primed Polymorphonuclear Leukocyte CD11b Induces Apoptosis and Prevents Cell Activation.

Heparin is known to have anti-inflammatory effects, yet the mechanisms are not completely understood. In this study, we tested the hypothesis that heparin has a direct effect on activated polymorphonuclear leukocytes (PMNLs), changing their activation state, and can explain its anti-inflammatory effect. To test our hypothesis, we designed both in vitro and ex vivo studies to elucidate the mechanism by which heparin modulates PMNL functions and therefore the inflammatory response. We specifically tested the hypothesis that priming of PMNLs renders them more susceptible to heparin. Amplified levels of CD11b and increased rate of superoxide release manifested PMNL priming. Increase in cell priming resulted in a dose-dependent increase in heparin binding to PMNLs followed by augmented apoptosis. Blocking antibodies to CD11b inhibited heparin binding and abolished the apoptotic response. Moreover, heparin caused a significant dose-dependent decrease in the rate of superoxide release from PMNLs, which was blunted by blocking antibodies to CD11b. Altogether, this study shows that the interaction of heparin with the PMNL CD11b results in cell apoptosis and explains heparin's anti-inflammatory effects

Massive Transfusion in Wartime: Experience from Northern Israel, Summer 2006.

Abstract In July 2006 hostilities erupted in Israel/Lebanon. Reported here is the experience of three medical centers in Northern Israel during 33 days of the warfare; the Rambam Health Care Campus in Haifa - a level I trauma center, the Rebecca Sieff Hospital in Safed and the Western Galilee Hospital in Nahariah - both secondary trauma centers. 504, 1138 and 868 wounded were presented to the three medical centers and 281, 415 and 195, respectively, required hospitalization. Sixty, 32 and 15 hospitalized patients were concomitantly transfused in each corresponding center, representing 20%, 7% and 7%, respectively, of admitted patients. Patients with an injury severity score of ≥16 had a higher need for blood products than those less severely injured, with a mean packed red blood cell (PRBC) transfusion of 7 versus 4 units (p=0.03) and FFP transfusion of 13 versus 1.5 units (p=0.002), respectively. Twenty four soldiers and one civilian had massive transfusions and twenty three of these patients survived. The median ratio between transfused fresh frozen plasma (FFP) and packed red blood cells (PRBC) was 0.8, ranging from a ratio of 0.25 to 1.3. Among 25 massively transfused patients 21 received cryoprecipitate and 19 - platelets. The median prothrombin time (INR) and partial thromboplastin time (PTT) increased during the first 2 hours after admission from 1.29 to 1.51 and from 33.6 seconds to 39 seconds, respectively. In the cohort of massively transfused patients 3 individuals additionally received 3 g of tranexamic acid, while another 2 patients were treated with recombinant factor VII. In conclusion, massively transfused patients with wartime penetrating injuries have an ongoing coagulopathy despite vigorous replacement therapy, which needs to be continued until the patients are stabilized. Early intervention and consultation in the Emergency Room by transfusion-service specialists is essential to the overall management of critically and massively wounded patients in wartime. Wounded (hospitalized) Transfused patients Packed RBC units FFP units Cryo units Platelet units Massive transfusion (patients) Rambam 504 (281) 60 463 413 266 258 21 Rebecca Sieff 1138 (415) 32 134 34 50 30 4 Western Galilee 868 (195) 15 71 68 51 10 1</jats:p

The Role of Alpha 2 Macroglobulin in Chronic Activation of the Complement System in Patients with Chronic Lymphocytic Leukemia

The immunotherapy treatments offered for Chronic lymphocytic leukemia (CLL) activate various cellular and biochemical mechanisms, including the complement system. Recently it was shown that the classical complement pathway (CP) in CLL patients is persistently activated at a low level through (Michelis et al. PlosOne, 2019), and that the mechanism of chronic CP activation involves the formation of IgG-hexamers (Michelis et al. PlosOne, 2020). According to recent studies, formation of ordered IgG-hexamers occurs on cell surfaces via specific interactions between Fc regions of the IgG monomers, which occur only after antigen binding. The study investigated the formation of IgG-hexamers in CLL patients and normal controls (NC), their ability to activate complement, their incidence as cell-free and cell-bound forms and the identity of the antigen causing their formation. Sera from 30 patients and NC were used for separation of IgG-hexamers. The obtained IgG-hexamers were measure and used for assessment of CP activity. For evaluation of the presence of IgG-hexamers on blood cells, whole blood samples were stained and assessed by flow cytometry. Serum levels of IgG-hexamers were higher in patients compared to NC. Also, they activated the complement system to a higher extent in patients than in NC. Alpha 2 macroglobulin (A2M) was recognized as the antigen causing the hexamerization of IgG, and was found to be part of the hexamer structure by mass spectrometry, Western blot and flow cytometry analysis. The presence of A2M-IgG-hexamers on B-cells suggests that it may be formed on B cells surface and then be detached to become cell-free. Alternatively, it may form in the plasma and then attach to the surface of the cells. The exact course of A2M-IgG-hexamers formation in CLL should be further studied. The results in this study may be useful for improvement of current immunotherapy regimens. Disclosures Tadmor: Neopharm: Consultancy, Speakers Bureau; AbbVie: Consultancy, Speakers Bureau; Janssen: Consultancy, Speakers Bureau; Takeda: Consultancy, Speakers Bureau; Sanofi: Consultancy, Speakers Bureau; Medison: Consultancy, Speakers Bureau; 6. Novartis Israel Ltd., a company wholly owned by Novartis Pharma AG: Consultancy, Speakers Bureau. Aviv:Abbvie: Honoraria. </jats:sec

The Role of Alpha 2 Macroglobulin in IgG-Aggregation and Chronic Activation of the Complement System in Patients With Chronic Lymphocytic Leukemia

Chronic lymphocytic leukemia (CLL) is the most common leukemia in adults in the western world. One of the treatments offered for CLL is immunotherapy. These treatments activate various cellular and biochemical mechanisms, using the complement system. Recently it was shown that the complement system in CLL patients is persistently activated at a low level through the classical pathway (CP). The mechanism of chronic CP activation involves the formation of IgG-hexamers (IgG-aggregates). According to recent studies, formation of ordered IgG-hexamers occurs on cell surfaces via specific interactions between Fc regions of the IgG monomers, which occur after antigen binding. The present study investigated the formation of IgG-hexamers in CLL patients and normal (non-malignant) controls (NC), their ability to activate complement, their incidence as cell-free and cell-bound forms and the identity of the antigen causing their formation. Sera from 30 patients and 12 NC were used for separation of IgG- aggregates. The obtained IgG- aggregates were measured and used for assessment of CP activation. For evaluation of the presence of IgG- aggregates on blood cells, whole blood samples were stained and assessed by flow cytometry. Serum levels of IgG- aggregates were higher in CLL and they activated the complement system to a higher extent than in NC. Alpha 2 macroglobulin (A2M) was identified as the antigen causing the hexamerization/aggregation of IgG, and was found to be part of the hexamer structure by mass spectrometry, Western blot and flow cytometry analysis. The presence of A2M-IgG-hexamers on B-cells suggests that it may be formed on B cells surface and then be detached to become cell-free. Alternatively, it may form in the plasma and then attach to the cell surface. The exact time course of A2M-IgG-hexamers formation in CLL should be further studied. The results in this study may be useful for improvement of current immunotherapy regimens.</jats:p